Recombinant HIV type 1 (MJ412.W0M.ENV.C1) Glycoprotein 120 (a.a. 32-513) [His]

The N332 high-mannose glycan on the HIV-1 gp120 V3-loop is the target of many bNAbs. About 17% HIV isolates carry the N332 to N334 mutation, but the antibody recognition of the N334 N-glycan and its immunogenicity are not well characterized. Here we report the chemoenzymatic synthesis, antigenicity, and immunogenicity of the V3 N334 glycopeptides from HIV-1 A244 gp120, a key component in the partially successful Thai clinical trials. We found that synthetic V3 glycopeptide carrying a N334 high-mannose glycan could be recognized by bNAb PGT128 and PGT126 but not by 10-1074. Rabbit immunization with the synthetic three-component A244 glycopeptide immunogen elicited substantial glycan-dependent antibodies with broad reactivity. to various HIV-1 gp120/gp140 carrying N332 or N334 glycosylation sites. These results indicated that the N334 site is vulnerable and the A244 V3 glycopeptide represents a valuable immunogen for further HIV-1 vaccine studies.

The aims of this study were twofold. The first was to determine if human immunodeficiency virus (HIV)-1 glycoprotein (gp) 120 and transactivator of transcription (Tat) stimulate the release of endothelial microvesicles (EMVs). The second was to determine whether viral protein-induced EMVs are deleterious to endothelial cell function (inducing endothelial cell inflammation, oxidative stress, senescence and increasing apoptotic susceptibility). Human aortic endothelial cells (HAECs) were treated with recombinant HIV-1 proteins Bal gp120 (R5), Lay gp120 (X4), or Tat. EMVs released in response to each viral protein were isolated and quantified. Fresh HAECs were treated with EMVs generated under control conditions and from each of the viral protein conditions for 24 h. EMV release was higher (P < 0.05) in HAECs treated with R5 (141 +/- 21 MV/mu l), X4 (132 +/- 20 MV/mu l), and Tat (130 +/- 20 MV/ mu l) compared with control (61 +/- 13 MV/mu l). Viral protein EMVs induced significantly higher endothelial cell release of proinflammatory cytokines and expression of cell adhesion molecules than control. Reactive oxygen species production was more pronounced (P < 0.05) in the R5-, X4- and Tat-EMV-treated cells. In addition, viral proteinstimulated EMVs significantly augmented endothelial cell senescence and apoptotic susceptibility. Concomitant with these functional changes. viral protein-stimulated EMVs disrupted cell expression of micro-RNAs 34a, 126, 146a, 181b, 221, and miR-Let-7a (P < 0.05). These results demonstrate that HIV-1 gp120 and Tat stimulate microvesicle release from endothelial cells, and these microvesicles confer pathological effects on endothelial cells by inducing inflammation. oxidative stress, and senescence as well as enhancing susceptibility to apoptosis. Viral protein-generated EMVs may contribute to the increased risk of vascular disease in patients with HIV-1. NEW & NOTEWORTHY Human immunodeficiency virus (HIV)1-related proteins glycoprotein (gp) 120 and transactivator of transcription (Tat)-mediated endothelial damage and dysfunction are poorly understood. Endothelial microvesicles (EMVs) serve as indicators and potent mediators of endothelial dysfunction. In the present study we determined if HIV-1 R5- and X4-tropic gp120 and Tat stimulate EMV release in vitro and if viral protein-induced EMVs are deleterious to endothelial cell function. gp120 and Tat induced a marked increase in EMV release. Viral protein-induced EMVs significantly increased endothelial cell inflammation, oxidative stress, senescence, and apoptotic susceptibility in vitro. gpl 20- and Tat-derived EMVs promote a proinflammatory, pro-oxidative. prosenescent, and proapoptotic endothelial phenotype and may contribute to the endothelial damage and dysfunction associated with gp120 and Tat.